CpG motifs of bacterial DNA exert protective effects in mouse models of IBD by antigen-independent tolerance induction.
OBJECTIVE
Prophylactic treatment of mice with CpG motifs of bacterial DNA protects from experimental inflammatory bowel disease, at least partly via induction of inhibitory T-cells. The aim of this study was to elucidate whether these CpG-dependent protective effects require presence of bacterial flora suggesting antigen-specific regulatory activity.


METHODS
Germ-free BALB/c and IL-10(-/-) mice were treated with CpG-oligodeoxynucleotides (ODN), control-ODN, or PBS. CD4(+)CD62L(+) cells of these mice were transferred into SCID recipients. CpG-ODN-treated germ-free IL-10(-/-) mice were transferred into colitogenic environment. Monoclonal antibodies were used to neutralize TGF-beta and IFN-alpha/beta during CpG-ODN treatment. CD4(+)CD62L(+) cells of donors were evaluated for cytokine secretion and FOXP3, PD-1, and CD25 expression.


RESULTS
Compared to PBS or control-ODN treatment, CpG-ODN application to germ-free donors led to decreased intestinal inflammation as indicated by histology, decreased proinflammatory cytokines, and increased IL-10 secretion. Protection was also observed after cotransfer of cells from PBS and CpG-ODN treated donors. Anti-TGF-beta and anti-INF-alpha/beta partly reversed the protective CpG-ODN effect. CpG-ODN-treated germ-free IL-10(-/-) mice transferred into colitogenic environment developed significantly less colitis than controls but not recipients of IL-10(-/-)CD4(+)CD62L(+)cells. CD4(+)CD62L(+)cells of CpG-treated germ-free animals displayed increased expression of regulatory markers.


CONCLUSIONS
Even without pre-existence of bacterial flora CpG-ODN exposition induces tolerance, indicating that CpG-ODN-induced regulatory T-cells are not bacterial antigen specific. TGF-beta and IFN-alpha/beta play major roles in induction of regulatory cells, and although IL10-independent mechanisms play a role in CpG-ODN protection, this cytokine likely is important for the effector mechanism of CpG-ODN-induced regulatory T-cells.
Bleich
Andre
A
Institute for Laboratory Animal Science and Central Animal Facility, Hannover Medical School, Hannover, Germany. bleich.andre@mh-hannover.de
Janus
Lydia M
LM
Smoczek
Anna
A
Westendorf
Astrid M
AM
Strauch
Ulrike
U
Mähler
Michael
M
Hedrich
Hans-J
HJ
Fichtner-Feigl
Stefan
S
Schölmerich
Jürgen
J
Falk
Werner
W
Hofmann
Claudia
C
Obermeier
Florian
F
D019264:Adoptive Transfer; D000818:Animals; D000942:Antigens, Bacterial / Q000502:physiology*; D016207:Cytokines / Q000096:biosynthesis; D005856:Germ-Free Life; D007108:Immune Tolerance; D015212:Inflammatory Bowel Diseases / Q000517:prevention & control*; D016753:Interleukin-10 / Q000502:physiology; D019041:L-Selectin / Q000032:analysis; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D009838:Oligodeoxyribonucleotides / Q000494:pharmacology*; D050378:T-Lymphocytes, Regulatory / Q000502:physiology; D016212:Transforming Growth Factor beta / Q000502:physiology
D000942:Antigens, Bacterial; C408982:CPG-oligonucleotide; D016207:Cytokines; D009838:Oligodeoxyribonucleotides; D016212:Transforming Growth Factor beta; D019041:L-Selectin; D016753:Interleukin-10